Production and characterization of guinea pig il-5 in baculovirus
-infected insect cells.
Mansour, Mahmoud, Michael Karmilowicz, Steven J. Hawrylik, Barbara
Nalcerio, Jane Angilly, Maryrose J. Conklyn, Craig M. Lilly, Jeffery
M. Drazen, S. E. Lee, David D. Auperin, Jeffrey R. Dewet, Victoria L.
Cohan, Henry J. Showell, and Dennis E. Danley.
Departments of Molecular Sciences and Immunology and Infectious
Disease, Pfizer Central Research Division, Groton, CT 06340 and
Department of Medicine, Brigham and Womens Hospital and Harvard
Medical School, Boston, MA 02115
APStracts 3:0021L, 1996.
To study the role IL-5 may play in altering airway function in asthma,
we have produced recombinant protein for exogenous administration to
guinea pigs. The guinea pig IL-5 (gpIL-5) cDNA was cloned by PCR
amplification of guinea pig spleen RNA and expressed as a secretion
product from recombinant baculovirus-infected Sf9 insect cell
cultures. The protein was purified to homogeneity by a four step
procedure which included immunoaffinity chromatography using
polyclonal antipeptide antibodies against a region of the mature
secreted cytokine. The cytokine was properly processed after the
signal sequence by the Sf9 cells, was glycosylated with terminal
mannose-containing oligosaccharide and had proper disulfide-linked
dimer structure as determined by SDS-PAGE. The purified preparation
was active in vitro and in vivo as determined by its ability to prime
human basophils to release leukotrine (LT)C4 in the presence of C5a
and to induce airway eosinophilia in naive guinea pigs.
Received 19 June 1995; accepted in final form 15 January 1996.
APS Manuscript Number L191-5.
Article publication pending Am. J. Physiol. (Lung Cell. Mol.
Physiology).
ISSN 1080-4757 Copyright 1996 The American Physiological Society.
Published in APStracts on 8 February 96